PROJECT SUMMARY/ABSTRACT The crisis of opioid overdose deaths and the rise in opioid use disorder (OUD) have crystalized the need to deepen our neurobiological understanding of OUD and identify novel therapeutic targets for this chronic medical disorder. Vulnerability to drug-related cues previously associated with opioid-taking behavior (?cue reactivity?) is thought to trigger cycles of opioid-seeking and relapse that contribute to high recidivism rates in OUD. Thus, an improved understanding of the mechanisms underlying opioid cue reactivity is crucial to generate new pharmacotherapeutic strategies to prevent relapse in OUD. Studies indicate that the medial prefrontal cortex (mPFC) is a crucial neural node involved in the mechanisms through which opioid-associated cues gain incentive-motivational properties that drive relapse during drug abstinence. The growth hormone secretagogue receptor 1? (GHS1?R) has been recently implicated in a number of reward processes as well as drug seeking behaviors, and we present exciting new data indicating that systemic pharmacological blockade of this receptor suppresses oxycodone cue reactivity in a rodent model. However, comprehensive data regarding the expression of this receptor in the brain is lacking, and the site of its action to attenuate oxycodone cue reactivity is unknown. Our preliminary data indicate enriched expression of the GHS1?R in the mPFC and propose that GHS1?R resident in mPFC neurons is a critical mediator of opioid relapse vulnerability. Through biochemical, behavioral, pharmacological, and genetic methodologies, the present study will address this hypothesis through two Specific Aims. Our first aim is to establish the mPFC neural framework for GHS1?R control of oxycodone cue reactivity via capillary electrophoresis-based immunoblot assays and immunohistochemistry. Our second aim is to interrogate the role of GHS1?R in mPFC in oxycodone cue reactivity via both intra-mPFC microinfusion of a clinically relevant GHS1?R inverse agonist as well as viral vector-mediated mPFC GHS1?R knockdown. Completion of the aims outlined in this proposal will provide the applicant with training in neurobiology and substance use disorders, experimental design, analysis and interpretation, critical thinking skills, as well as the proper implementation of valid statistical analyses. The outcomes of these studies will have a sustained, powerful impact on our field by identifying a key regulatory role for the GHS1?R in the mPFC in neural processes underlying cue reactivity, which will critically advance efforts to decrease relapse vulnerability and improve treatment outcomes in OUD.